转谷氨酰胺酶2抑制H1亚型流感病毒在MDCK细胞中的增殖

Transglutaminase 2 inhibits the proliferation of H1 subtype influenza virus in MDCK cells

摘要:

组织转谷酰胺酶(transglutaminase 2,TGM2)是一种普遍存在的多功能蛋白，与不同细胞的粘附和肿瘤形成有关。有证据表明，TGM2参与了宿主细胞与病毒间的相互作用，但是对于流感病毒在细胞内增殖的影响还未有报道。为了探究MDCK细胞中TGM2对H1N1亚型流感病毒增殖的影响，本研究构建了TGM2过表达和敲除的MDCK稳定细胞系，感染H1N1亚型流感病毒48 h后检测病毒NP和NS1蛋白mRNA和蛋白表达水平的变化，测定病毒滴度。结果显示，过表达TGM2能够有效抑制H1N1亚型流感病毒的NP、NS1基因的表达，而敲除TGM2可上调病毒NP、NS1基因的表达，其中NP蛋白的蛋白表达情况与mRNA水平一致。病毒增殖曲线结果显示，TGM2过表达后H1N1亚型流感病毒滴度显著降低，而敲除TGM2后结果相反，48 h时病毒滴度在敲除细胞中达到高峰，进一步证明TGM2在MDCK细胞中参与对H1N1亚型流感病毒增殖的抑制。通过对流感病毒应答信号途径下游基因的表达分析发现，TGM2可能通过促进激活JAK-STAT分子途径和抑制RIG-1信号途径，抑制H1N1亚型流感病毒增殖。以上发现对于揭示宿主细胞与病毒互作机制以及建立高产的流感疫苗生产基因工程细胞系具有重要意义。

Abstract:

Transglutaminase 2(TGM2) is a ubiquitous multifunctional protein,which is related to the adhesion of different cells and tumor formation.Previous studies found that TGM2 is involved in the interaction between host cells and viruses,but the effect of TGM2 on the proliferation of influenza virus in cells has not been reported.To explore the effect of TGM2 during H1N1 subtype influenza virus infection,a stable MDCK cell line with TGM2 overexpression and a knockout cell line were constructed.The mRNA and protein expression levels of NP and NS1 as well as the virus titer were measured at 48 hours after pot-infection with H1N1 subtype influenza virus.The results showed that overexpression of TGM2 effectively inhibited the expression of NP and NS1 genes of H1N1 subtype influenza virus,while knockout of TGM2 up-regulated the expression of the NP and NS1 genes,and the expression of the NP at protein level was consistent with that at mRNA level.Virus proliferation curve showed that the titer of H1N1 subtype influenza virus decreased significantly upon TGM2 overexpression.On the contrary,the virus titer in TGM2 knockout cells reached the peak at 48 h,which further proved that TGM2 was involved in the inhibition of H1N1 subtype influenza virus proliferation in MDCK cells.By analyzing the expression of genes downstream of influenza virus response signaling pathway,we found that TGM2 may inhibit the proliferation of H1N1 subtype influenza virus by promoting the activation of JAK-STAT molecular pathway and inhibiting RIG-1 signaling pathway.The above findings are of great significance for revealing the mechanism underlying the interactions between host cells and virus and establishing a genetically engineering cell line for high-yield influenza vaccine production of influenza virus.